Project Summary Successful DNA replication is essential to maintain genome integrity and prevent disease. In human cells, over 6 billion base pairs of DNA need to be replicated accurately and packaged into chromatin each cell division cycle. Furthermore, these processes must occur in the context of defects in the DNA template such as DNA damage, which present challenges to both genetic and epigenetic inheritance. The replisome is a highly regulated, dynamic machine that must work with speed and precision to duplicate the chromosomes successfully. Failures during DNA replication cause mutations, epigenetic changes, and other chromosomal aberrations that ultimately cause disease such as cancer. Our guiding hypothesis is that replication stress induces a series of DNA damage responses (DDR) that promote fork stabilization, repair, and recovery, allowing the completion of genome and chromatin replication. In this project we aim to define mechanisms that promote DNA replication especially in the context of genotoxic stress. The project uses innovative methodologies including a technology we developed called iPOND to isolate proteins on nascent DNA. Completion of this project will define DNA and chromatin replication, repair, and signaling mechanisms that act to ensure DNA replication is completed faithfully.